According to the U.S. Dept. of Health and Human Services, acetaldehyde is "reasonably anticipated to be a human carcinogen". When administered to laboratory animals by inhalation, acetaldehyde produces nasal and laryngeal carcinomas. Acetaldehyde occurs widely in the human environment, is a major constituent of cigarette smoke, and is the main metabolite of ethanol. Levels of acetaldehyde in the environment may increase with the introduction of ethanol-containing fuels. DNA adducts are critical in the carcinogenic process. In this program, we have identified DNA adducts of acetaldehyde including the major adduct N2- ethylidene-dGuo (adduct 1), the 1,A/2-propano-dGuo adducts 3 which are also formed from crotonaldehyde, and a related interstrand cross-link (adduct 4). We have developed highly sensitive mass spectrometric methods to quantify adducts 1 and 3 in human tissues. We have demonstrated that adduct 1 can be quantified in low microgram amounts of DNA, that it is an endogenous DNA adduct, and that its levels are influenced by cigarette smoking. We have also shown that adduct 3 is present in human lung DNA and has miscoding potential in human cells. In this renewal application, we propose to continue our studies on acetaldehyde DNA adducts to test our overall hypothesis that they are involved as causes of human cancer, particularly of the lung, and head and neck. Our goal is to investigate the occurrence in humans and the biological significance of acetaldehyde DNA adducts. Our specific aims are: 1. Quantify levels of adducts 1, 3, and the related acrolein-derived adduct 5 in human lung DNA from current smokers (confirmed by urinary cotinine) and compare levels of these adducts to those derived from benzo[a]pyrene (BaP) and tobacco-specific nitrosamines in the same tissues. 2a. Determine the influence of polymorphisms in alcohol dehydrogenase (ADH1C) and aldehyde dehydrogenase (ALDH2) genes on levels of adduct 1 in leukocyte DNA of non-smokers who consume alcohol. b. Quantify levels of adduct 1 in leukocyte DNA of non-smoking, non-drinking Chinese women who regularly engage in wok cooking compared to those who do not. 3. Investigate the genotoxic properties of adduct 1 in cells, and by studies in A/J mice which compare adduct formation and tumorigenicity of compounds that do or do not generate acetaldehyde. 4. Investigate the mechanism of translesion synthesis across adduct 3 and "half-excised" interstrand cross-links. We have shown that these adducts are bypassed by mammalian DNA polymerases in cells. Here we propose to identify those polymerases and characterize the translesion synthesis by in vitro and cellular approaches. These studies will provide critical data on the occurrence and biological significance of acetaldehyde DNA adducts and their possible role in human cancer.